Interaction of 125I-labelled ceruloplasmin with human trophoblast cells in vitro.
In vitro binding, internalization and release of the plasma protein ceruloplasmin was investigated using primary cultures of human placental trophoblast cells. Binding of 125I-labelled ceruloplasmin at 4 degrees C reached equilibrium by 5-6 h; binding was linear throughout all concentrations tested (1 nM-3.3 microM). Addition of greater than 5-10 microM unlabelled ceruloplasmin or a variety of other proteins (albumin, transferrin, IgG) were equally effective in displacing bound ceruloplasmin in a concentration-dependent manner. When cells were incubated at 37 degrees C, the majority of surface-bound 125I-labelled ceruloplasmin was released directly to the extracellular medium. Trypsin-resistant radioactivity increased to 18 per cent of initially bound ceruloplasmin within 1 min, declining to 5 per cent by 2 h. The acquisition of trypsin-resistant radioactivity was unaffected by the addition of a variety of metabolic inhibitors and no evidence of intracellular degradation of ceruloplasmin was found. In summary, our results suggest that the majority of ceruloplasmin binding to trophoblast cells is nonspecific, of low affinity, and easily dissociable at 4 degrees C. Only a small amount of ceruloplasmin appeared to be internalized, by an as yet unknown mechanism.
Kennedy
M L
ML
Department of Human Anatomy, School of Medicine, University of California, Davis 95616.
Douglas
G C
GC
King
B F
BF
Lonnerdal
B
B
D000643:Ammonium Chloride / Q000494:pharmacology; D001386:Azides / Q000494:pharmacology; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002570:Ceruloplasmin / Q000493:pharmacokinetics*; D003513:Cycloheximide / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005461:Fluorine / Q000494:pharmacology; D006801:Humans; D007074:Immunoglobulin G / Q000494:pharmacology; D066298:In Vitro Techniques; D008985:Monensin / Q000494:pharmacology; D012709:Serum Albumin / Q000494:pharmacology; D014168:Transferrin / Q000494:pharmacology; D014327:Trophoblasts / Q000378:metabolism*
D001386:Azides; D007074:Immunoglobulin G; D012709:Serum Albumin; D014168:Transferrin; D000643:Ammonium Chloride; D005461:Fluorine; D008985:Monensin; D003513:Cycloheximide; D002570:Ceruloplasmin